The study of cellular membrane-associated proteins and membrane-bound proteins has been hampered by the need to provide unique microenvironments in which such proteins may exist naturally in the cell or as extracellular components. Much study and effort has been applied to this problem because the cellular components, upon isolation without membrane or lipid components to help solubilize them, form intractable and insoluble proteinaceous aggregates that precipitate out of solution in most simple aqueous environments. It has been found that such membrane proteins and components must not be isolated to absolute purity, but rather isolated such that they are transferred from the in vivo environment to an in vitro environment comprised of purified, synthetic lipids specially formulated and suited to provide the targeted protein or component with an physical and chemical environment much like that from which it is derived in vivo. The technique of purifying membrane proteins and components in the context of synthetic purified lipid or lipid-like molecules often provides useful quantities of such proteins and components for further characterization and use.
There exists several different families or classes of lipid or lipid-like molecules that help facilitate the purification and study of membrane proteins and components. In the past, purified preparations of naturally occurring lipids have been used, as well as synthetic lipids. Lipid-like molecules have also found much use in this field, including amphiphiles made of peptides and other chemical components which possess amphiphilic properties.
However, there still exists a great need for continued expansion of the field of membrane protein study to provide additional chemistries and lipid, or lipid-like, components to researchers, scientists, and industry for purifying proteins and components that, despite all available techniques, remain intractable and unable to be characterized because the proper chemistries and biochemistries have yet to be found which provide the exact unique microenvironment ideally suited for the isolation of those specific target proteins and/or component. To this end, the present inventors have endeavored to expand the field of membrane biology by finding and developing additional biochemical molecules having lipid-like characteristics which, when brought together under the proper conditions, form a multitude of different and expansive methodologies for the purification, isolation, and study of those membrane-associated or membrane-bound proteins and/or biochemical components which have thus far resisted purification or isolation and therefore, resisted further study.
Membrane-bound or membrane-associated proteins and/or biochemical components have been found to be highly useful in the study of human diseases and treatment thereof. Many neurological disorders may be traced directly to causalities that arise from mutations or other malfunctions, such as under-expression or over-expression, of membrane-bound or membrane-associated proteins or other biochemical components. Transmembrane proteins have been found to play crucial biological roles in intracellular communication and signaling, intracellular communication for instance between organelles and cytosol, ion transport, extracellular matrix interaction, tissue and arterial health and viral susceptibility. (See, for instance, Cobbold et al., “Aberrant trafficking of transmembrane proteins in human disease,” Trends Cell Biol., 13(12):639-647, 2003). Transmembrane proteins play key roles in diabetes, hypertension, depression, arthritis, cancer and neurological diseases such as cystic fibrosis. For instance cystic fibrosis has been tightly linked to the function of a transmembrane protein called Cystic Fibrosis Transmembrane Receptor (CFTR). A transmembrane form of the prion protein has been linked to neurodegenerative diseases. (See, Hegde et al., Science, 279(5352):827-834, 1998). Several nuclear envelope transmembrane proteins have been associated with signaling functions at the nuclear envelope which is involved in human diseases affecting skeletal muscle development. (See, Chen et al., BMC Cell Biology, 7:38, 2006).
The pharmaceutical industry has particularly benefited from the study of such membrane structures. It is estimated that about half of all potential pharmaceutical targets are membrane proteins such as ion channels and G-protein coupled receptors (GPCRs). The pharmaceutical and biotechnology industry have been able to produce and isolate sufficient quantities of a small number of membrane proteins and components to enable characterization of these targets, allowing production of biologics and pharmaceuticals useful in the treatment and prevention of diseases linked to these targets. Furthermore, use of such lipid and lipid-like chemistries has allowed the advancement of such fields as chemotherapy and virology. For instance, modern vaccines have greatly benefited from the development of modern and industrially useful lipid preparations which are highly efficient in triggering precise immunological responses in animals and humans. (See, for instance, Copland et al., “Lipid based particulate formulations for the delivery of antigen,” 1 mm. Cell Biol., 83:97-105, 2005). The field of chemotherapy has highly benefited from progressive research performed on specific pharmaceutical emulsions which allow precise targeting and localization of otherwise very poisonous and toxic substances to only cancerous tissues and organs, preventing damage to other healthy tissue. (See, Kishor M. Wasan, “Role of Lipid Excipients in Modifying Oral and Parenteral Drug Delivery: Basic Principles and Biological Examples,” John Wiley & Sons, Inc., Hoboken, N.J., 2007; and Davis et al., “Lipid Emulsions as Drug Delivery Systems,” Annals of the New York Acad. Of Sciences, 507:75-88, December 1987).
A wide variety of lipid and lipid-like molecules, also called surfactants, are disclosed in the literature of various polarities, sizes, hydrophobicities, and the like. Lipid science has advanced to provide a myriad number of variations on sugar-like lipids, phospholipids with polar head groups, lipids with multiple polar head groups, polar head groups varying in polarity, and carbohydrate chains varying in size, composition and properties. Presently provided is a class of carbohydrate surfactants having additional chemical functionality. The lipid derivatives of the carbohydrates provided here, such as, but not limited to, maltoside-based alkyne and azide derivatives, as well as phosphocholine derivatives. The chemical functionalities imparted to these derivatives may be useful in conjugating them to other groups. The conjugated surfactants may then provide interesting new molecules having unique biochemical and biophysical properties which heretofore have not been available or easily accessible to the modern chemist or biochemist. These novel properties and characteristics provide industry and academia with an expanded arsenal of as yet untested biochemical surfactants, potentially possessing a wide variety of biochemical and biophysical characteristics, which when applied to the study of lipid-bound, or membrane-associated proteins and/or other biochemical components, may yield unprecedented results. This expanded repertoire of combination surfactants may therefore lead to new treatments and perhaps prevention of diseases that have as yet remained intractable due to the nature of the membrane- or lipid-related target to which the disease may be linked.
Among other aspects, the present invention provides compositions of surfactant derivatives and the like, methods of making the same, and methods of using the same, for instance in the formation of micelles and micelle-like structures, that address the above noted needs. A complete understanding of the invention will be obtained upon review of the following.